1. Field of the Invention
This invention relates generally to gene expression in normal and neoplastic cells, and specifically to a novel tumor suppressor gene, HIC-1, and its gene product.
2. Description of Related Art
Advances in recombinant DNA technology have led to the discovery of normal cellular genes such as proto-oncogenes and tumor suppressor genes, which control growth, development, and differentiation. Under certain circumstances, regulation of these genes is altered and they cause normal cells to assume neoplastic growth behavior. There are over 40 known proto-oncogenes and tumor suppressor genes to date, which fall into various categories depending on their functional characteristics. These include, (1) growth factors and growth factor receptors, (2) messengers of intracellular signal transduction pathways, for example, between the cytoplasm and the nucleus, and (3) regulatory proteins which influence gene expression and DNA replication (e.g., transcription factors).
Chromosome 17p is frequently altered in human cancers, and allelic losses often coincide with mutations in the p53 gene at 17p13.1 (Vogelstein, B., et al., Cell, 70:523, 1992). This gene is one of the most frequently altered tumor suppressor genes in human neoplasms. However, in some tumor types, 17p allelic loss occurs at a high frequency in regions distal to p53 and in the absence of p53 mutations. For instance, 60% of breast cancers lose 17p alleles while only 30% of these tumors contain p53 mutations (Chen, L-C., et al., Proc. Natl. Acad. Sci. USA, 88:3847, 1991; Takita, K., et al., Cancer Res., 52:3914, 1992; Deng, G., et al, Cancer Res., 54:499, 1994; Cornelis, R. S., et al., Cancer Res., 54:4200, 1994). Furthermore, in one study of breast cancer, the independent loss of 17p13.3 alleles was accompanied by increased levels of p53 MRNA.
Human cancer cells typically contain somatically altered genomes, characterized by mutation, amplification, or deletion of critical genes. In addition, the DNA template from human cancer cells often displays somatic changes in DNA methylation (E. R. Fearon, et al., Cell, 61:759, 1990; P. A. Jones, et al., Cancer Res., 46:461, 1986; R. Holliday, Science, 238:163, 1987; A. De Bustros, et al., Proc. Natl. Acad. Sci., USA, 85:5693, 1988); P. A. Jones, et al., Adv. Cancer Res., 54:1, 1990; S. B. Baylin, et al., Cancer Cells, 3:383, 1991; M. Makos, et al., Proc. Natl. Acad. Sci., USA, 89:1929, 1992; N. Ohtani-Fujita, et al., Oncogene, 8:1063, 1993). However, the precise role of abnormal DNA methylation in human tumorigenesis has not been established. DNA methylases transfer methyl groups from the universal methyl donor S-adenosyl methionine to specific sites on the DNA. Several biological functions have been attributed to the methylated bases in DNA. The most established biological function is the protection of the DNA from digestion by cognate restriction enzymes. The restriction modification phenomenon has, so far, been observed only in bacteria. Mammalian cells, however, possess a different methylase that exclusively methylates cytosine residues on the DNA, that are 5' neighbors of guanine (CpG). This methylation has been shown by several lines of evidence to play a role in gene activity, cell differentiation, tumorigenesis, X-chromosome inactivation, genomic imprinting and other major biological processes (Razin, A., H., and Riggs, R. D. eds. in DNA Methylation Biochemistry and Biological Significance, Springer-Verlag, New York, 1984).
A CpG rich region, or "CpG island", has recently been identified at 17p13.3, which is aberrantly hypermethylated in multiple common types of human cancers (Makos, M., et al., Proc. Natl. Acad. Sci. USA, 89:1929, 1992; Makos, M., et al., Cancer Res., 53:2715, 1993; Makos, M., et al, Cancer Res. 53:2719, 1993). This hypermethylation coincides with timing and frequency of 17p losses and p53 mutations in brain, colon, and renal cancers. Silenced gene transcription associated with hypermethylation of the normally unmethylated promoter region CpG islands has been implicated as an alternative mechanism to mutations of coding regions for inactivation of tumor suppressor genes (Baylin, S. B., et al., Cancer Cells, 3:383, 1991; Jones, P. A. and Buckley, J. D., Adv. Cancer Res., 54:1-23, 1990). This change has now been associated with the loss of expression of VHL, a renal cancer tumor suppressor gene on 3p (J. G. Hernan, et al., Proc. Natl. Acad. Sci. USA, 91:9700-9704, 1994), the estrogen receptor gene on 6q (Ottaviano, Y. L., et al., Cancer Res., 54:2552,1994) and the H19 gene on 11 p (Steenman, M. J. C., et al., Nature Genetics, 7:433, 1994).
For several human tumor types, a second tumor suppressor gene may reside distal to, and be interactive with, the p53 gene at chromosome 17p13.1. There is a need to identify tumor suppressor genes in order to develop the appropriate methodologies for increasing or decreasing their expression in cells where aberrant expression is observed. Through characterization of a 17p13.3 CpG island which is aberrantly hypermethylated in multiple common human tumor types, the present invention provides such a gene. HIC-1 (hypermethylated in cancer) is a novel zinc finger transcription factor gene which is ubiquitously expressed in normal tissues, but underexpressed in tumor cells (e.g., breast, lung, colon, fibroblasts) where it is hypermethylated. A p53 binding site is located in the 5' flanking region of HCI-1. Overexpression of a wild-type p53 gene in colon cancer cells containing only a mutant p53 allele, results in 20-fold activation of HIC-1 expression.
The present invention shows that many human cancers exhibit decreased HIC-1 expression relative to their tissues of origin. The limitation and failings of the prior art to provide meaningful markers which correlate with the presence of cell proliferative disorders, such as cancer, has created a need for markers which can be used diagnostically, prognostically, and therapeutically over the course of such disorders. The present invention fulfills such a need.